NERP015: The in vivo relevance and therapeutic applications of IFITIVI Proteins The interferon inducible transmembrane (IFITM) proteins are a family of recently-characterized interferonstimulable viral restriction factors that block entry by a receptor-independent mechanism. The IFITM proteins are highly effective against several NIAID Priority Pathogens including influenza A viruses (lAV), Marburg and Ebola viruses, the SARS coronavirus. West Nile virus, and dengue virus. The proposed project consists of two parallel aims. The first aim is to perform a comprehensive study of the in vivo effects of the IFITM proteins using Ifitm-knockout mouse models of lAV pneumonia. The second consists of an innovative small molecule screen that will identify lead therapeutic compounds whose activity mimics the effects of the IFITM proteins. The first aim, determination of in vivo function, is crucial to understanding the efficacy and mechanism of action of the IFITM proteins in the context of a living animal. Two Ifitm knockout mice are available as models for IFITM deletion. The first, the IfitmDel mouse, lacks all murine orthologs of the human IFITM genes relevant to viral restriction (IFITMs 1, 2, and 3). The second, the IfitmSegfp mouse, lacks only IFITM3, the most potent restriction factor in in vitro studies. We will infect these mice with respiratory-tropic lAV PR/8/34 and systemic lAV WSN/33 and measure IFITM-dependent differences in disease progression, viral loads, tissue tropism, cytokine responses, and adaptive immune responses. The second aim, identification of IFITM-like antiviral drugs, will search for therapeutics using a novel, internally-controlled, cell-based assay. The assay is based on infection of interferon-insensitive cells with pairs of IFITM-restricted and -unrestricted pseudoviral particles. Compounds will be selected that block entry of IFITM-restricted pseudoviruses but permit entry of unrestricted pseudoviruses. The inclusion of the unrestricted pseudoviral control will permit exclusion of compounds with non-specific and deleterious effects on cell physiology. At the same time, this approach will identify broad-spectrum therapeutics with efficacy against several Priority Pathogens.